Cell transplantation therapies using cell sheets have been recently studied as medicine to supplement tissue deficient in cells due to disease or injury with cells. In particular, cardiomyocyte replacement therapy has been studied because adult cardiomyocytes rarely proliferate and, therefore, loss of cardiomyocytes caused by ischemic heart disease etc. becomes irreversible damage.
Accordingly, use of cardiomyocyte sheets produced from cardiomyocytes has been proposed. For such regenerative therapies to be successful, sufficient engraftment of the cardiovascular cell fraction is essential. Moreover, methods using, as cardiomyocytes for such a purpose, cardiac cell fractions derived from embryonic stem cells (ESC) or induced pluripotent stem cells (iPSC) (Non-patent documents 1 and 2 and Patent document 1) have been studied.
Reduction of cardiac function in rat after myocardial infarction was confirmed to be suppressed by transplanting 3 layers cardiac tissue sheets produced from these cells. However, this reduction was not caused by direct effects of the induced cardiomyocytes, but by suppression of left ventricle remodeling by indirect paracrine effects including angiogenesis due to cytokines released by the cells (Non-patent document 3).
Therefore, administration of more cardiomyocytes is necessary to have cardiomyocyte sheets engrafted onto myocardial layers and, to that end, thicker sheets containing large amounts of cardiomyocytes are desired. However, it has been confirmed that even upon transplantation of 4 or more layers of cardiomyocyte sheets without blood vessels, some cells do not survive due to the shortage of oxygen and nutrient supply and therefore the number of engrafted cells does not change. Accordingly, further technological development is considered to be necessary to produce effective cardiomyocyte sheets in cell replacement therapies.
Gelatin hydrogel is a biodegradable biomaterial that can be used as a cell culture substrate or scaffolding material and inclusion of particles of the gel in aggregates of cultured cells, for example, has been reported to improve oxygen supply, etc., and thus the condition of cells (Non-patent document 4 and Patent document 2).
Known methods of using gelatin hydrogel to produce cell sheets for transplant include a method of attaching gelatin hydrogel to sheets to make the cell sheets easy to be detached from the cell culture substrate (Patent documents 3, 4, and 5).